This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. AurF, a novel oxygenase from S. thioluteus, catalyzes the unusual 6-electron oxidation of p- aminobenzoic acid (pABA) to p-nitrobenzoic acid (pNBA), an essential structural component of the antibiotic aureothin. We have demonstrated that AurF is a member of dinuclear non-heme- iron enzymes that are involved in numerous biological oxidation reactions (e.g. soluble methane monooxygenase). Using a combination of biochemical, kinetic, and spectroscopic techniques, we have identified a chemically competent peroxo-diiron(III/III) intermediate in this reaction. In the absence of substrate, pABA, this intermediate accumulates to nearly stoichiometric amounts, but it readily oxidizes the substrate. The peroxo-diiron(III/III) intermediate has markedly different spectroscopic parameters compared to other, well characterized peroxo-diiron(III/III) that are believed to have a ?-1,2-peroxo unit. Its optical transition (possibly a peroxide-to-iron charge transfer band) is observed around 500 nm, and it has a M[unreadable]ssbauer isomer shift of beta = 0.50 mm/s. ?-1,2-Peroxo-diiron(III/III) intermediates exhibit M[unreadable]ssbauer isomer shifts beta >0.60 mm/s and a LMCT band at ~700 nm.